Trypanosomatids are parasites that cause debilitating and often fatal diseases in humans and livestock. A particularly serious problem in sub-Saharan Africa is caused by Trypanosoma brucei, a parasite spread by the bite of an infected Tsetse fly. T. brucei causes disease by proliferating in the blood, tissue spaces, and eventually the central nervous system of its mammalian host. Related parasites are equally important to human populations in other parts of the world, including South and Central America, Southeastern Asia and the Middle East. Our long-term goal is to understand, in biochemical and genetic detail, the mechanisms that coordinate gene expression in trypanosomatids. To achieve this goal, we are specifically investigating the mechanism used by the parasites to regulate their mRNA expression pattern. mRNA expression gives rise to a characteristic and essential proteome and determines the metabolic capabilities of the parasites. As a pathway into understanding mRNA expression control, we are producing the mRNA-binding protein proteome for T. brucei procyclic parasites. We currently know of no other similar undertaking but are aware that this approach is extremely useful in the study of yeast biology and mammalian disease exploration. Our proposed two-year project is grounded in our experience in in vivo UV-crosslinking of protein-RNA complexes and analysis of two mRNA-binding proteins. The proposed project will establish the RNA-binding protein-mRNA proteome, in aim 1, and provide experimental verification of the dataset by analyzing the mRNA targets of two specific mRNA- binding proteins in aim 2.